Infection of cells either permissive or non-permissive for SV-40 lytic growth results in a stimulation of cellular DNA synthesis and an induction of many enzymes whose control is coordinated with the cell cycle. We propose to explore these known phenomenon further by isolating DNA sequences from cellular chromosomes which can serve as origins of DNA replication in vivo. The interaction of viral proteins with these sequences both in vivo and in-vitro will be investigated. Furthermore we wish to clone from human cells (via a novel route) the DNA sequences encoding for the enzyme thymidine kinase. This enzyme is induced in a viral infection and we will investigate via in-vivo and in-vitro experiments the nature of the induction. Finally we wish to examine the modes by which the "transformed" phenotype may evolve in culture and how this evolution is related to the initial type of selection imposed. These experiments include the use of novel forms of SV-40 molecules created by recombinant DNA techniques. We propose to transform a rat TK- cell line, possessing a characteristic "normal" growth phenotype, with DNA molecules containing both the TK gene and the SV40 early region. Selection for colonies will be based upon the positive requirement for the TK enzyme in HAT media. These clones will then be tested separately for their abilities to display the transformed phenotype by various criteria. We are particularly interested in the possible rearrangement (amplification or translocation) of the integrated viral genome coordinate with selection for its expression.